FIELD OF THE INVENTION

This invention relates to a method of extracting cannabinoids from plant waxes. In one aspect it relates to separating cannabinoids from waste waxes generated during conventional extraction processes.

BACKGROUND TO THE INVENTION

Cannabinoids are compounds that can interact with cannabinoid receptors of the endocannabinoid system in the body. At least 113 distinct cannabinoids have been isolated from the cannabis plant. Cannabinoids also occur in other plants and plant species such as rhododendron, licorice, liverwort, Echinacea (Echinacea purpurea and Echinacea angustifolia for example), Acmeiia oleracea and Helichrysum umbraculigerum. The most well-known cannabinoids are tetrahydrocannabinol (THC), cannabidiol (CBD) and cannabinol (CBN). Classical cannabinoids are structurally related to the most notable phytocannabinoid, THC. Cannabinoids are used for various purposes, including in medicine, for the treatment of neuropathic pain, spasticity and nausea due to chemotherapy. CBD is not psychoactive and is a popular natural remedy for various ailments. CBD extracts are used in medicines, dietary supplements, bath products, drinks and food.

Cannabinoids are concentrated in the viscous resin or waxes produced in plants. Cannabinoids can be separated from a plant by extraction with supercritical carbon dioxide or organic solvents. Hydrocarbons and alcohols are often used as solvents. Once extracted, isolated components can be separated using wiped film vacuum distillation or other distillation techniques.

Conventional extraction of cannabinoids from plants generates waste wax. Generally, cold extraction processes are used to extract medicinal cannabinoids from cannabis plants. The cannabinoids are extracted from the plant material into a solvent phase. However, the cannabinoids are not extracted in a pure form since several flavour compounds and waxes are co-extracted. To remove the waxes, the extraction solution is cooled to below -40 °C to precipitate the waxes in a step called winterisation. This step allows solidification of the waxes, which are then simply filtered out of the solution and removed from the processing stream. However, the waxes still entrap significant amounts of cannabinoids. The concentration of cannabinoids in the waste waxes may range between about 40 and 50 weight percentage (wt%) of the dry wax, for example. A current bottleneck in the cannabinoid processing industry is obtaining sufficient quantities of plant material from which to extract cannabinoids and in particular medical grade cannabinoids. Accordingly, there is a need for an effective method of separating or extracting cannabinoids from plant wax, particularly the extraction residues or waste wax from a primary extraction process to recover more cannabinoids.

The preceding discussion of the background to the invention is intended only to facilitate an understanding of the present invention. It should be appreciated that the discussion is not an acknowledgment or admission that any of the material referred to was part of the common general knowledge in the art as at the priority date of the application.

SUMMARY OF THE INVENTION

In accordance with the invention there is provided a method of extracting cannabinoids from plant waxes, the method comprising the steps of: dissolving the plant waxes in a selected solvent having a normal boiling point above 50 °C and a relative polarity in a range from about 0.1 to about 0.4 at an elevated temperature, thereby forming a solution of the plant waxes and the cannabinoids contained therein; and allowing the solution to cool to precipitate the plant waxes whilst the cannabinoids remain in solution.

The selected solvent may have a normal boiling point in a range from about 50 °C to about 1 10 °C. The selected solvent may have a relative polarity in a range from about 0.103 to about 0.353.

The solvent may be selected from the group consisting of 1 ,1 -dichloroethane, 1 ,2-dichloroethane, 2-butanone, 2-methyl tetrahydrofuran, 3-pentanone, benzene, chloroform, diethylamine, 1 ,2- dimethoxyethane, dioxane, ethyl acetate, methyl acetate, methyl tert-butyl ether, tetrahydrofuran, and any mixture thereof. The solvent may be selected from the group consisting of 1 ,1 - dichloroethane, 2-butanone, 2-methyl tetrahydrofuran, 3-pentanone, 1 ,2-dimethoxyethane, ethyl acetate, methyl acetate, methyl tert-butyl ether, and any mixture thereof.

The selected solvent may substantially lack hydrogen bond donation ability, so as to be substantially unreactive towards the cannabinoids and, optionally, the lipophilic compounds of the plant waxes under conditions of elevated temperature. The elevated temperature may be a temperature obtainable by mild heating. The elevated temperature may be a temperature in a range from about 45 °C to about 60 °C. The hydrogen bond donation ability may be quantified in terms of a Kamlet-Taft acidity parameter (a) approximating 0 (zero).

The solvent may be selected from the group consisting of 1 ,2-dimethoxyethane, 2-methyl tetrahydrofuran, 3-pentanone, ethyl acetate, methyl acetate, methyl tert-butyl ether, and any mixture thereof. The solvent may be ethyl acetate.

The solution may also be agitated during the heating step. Agitation may be by stirring.

The solution may be allowed to cool to below 30 °C to precipitate most of the plant waxes, preferably to about 20 °C to precipitate substantially all the plant waxes. Cooling may be by the removal of heat.

The method may further include the step of removing the precipitate from the solution. The removal of the precipitate may be by filtration or centrifugation, preferably filtration. The steps of the method may be repeated on the wax precipitate separated from the solution. The solvent may be recycled.

The plant waxes may be waste waxes obtained after an earlier or initial cannabinoid extraction process carried out on plants. The earlier or initial cannabinoid extraction process may be a cold cannabinoid extraction process involving a cooling step carried out on a first or primary extraction solution that results in the precipitation of waste waxes which include residual cannabinoids.

The cannabinoids extracted with the method described herein may be further processed into medical or food grade cannabinoids.

The plant waxes may be cannabis plant waxes.

In accordance with a second aspect of the invention, there is provided a cannabinoid extract obtained by the method as described above.

The cannabinoid extract may be substantially free of plant waxes. The cannabinoid extract includes at least about 15% of the original amount of cannabinoids in the plant waxes, preferably at least about 30%, more preferably at least about 60%.

Embodiments of the invention will now be described, by way of example only, with reference to the accompanying drawings. BRIEF DESCRIPTION OF THE DRAWINGS

In the drawings:

Figure 1 is a flow diagram that illustrates a method of extracting cannabinoids from plant waxes; and

Figure 2 is a flow diagram including process streams and operation units for an exemplary two-stage process for extracting cannabinoids from plant waxes.

DETAILED DESCRIPTION WITH REFERENCE TO THE DRAWINGS

Plant waxes are mostly found in an amorphous layer on the outer surface of plants but can also occur intracellularly. They are synthesized in the epidermal cells of most plants and exuded onto the surface. Plant waxes comprise a mixture of organic compounds that are lipophilic and malleable solids near ambient temperatures. More particularly, plant waxes consist of a mixture of substituted long-chain aliphatic hydrocarbons, containing alkanes, alkyl esters, fatty acids, primary and secondary alcohols, diols, ketones and aldehydes. The chemical composition of a particular mixture of plant waxes depends on the species of plant it is obtained from. Cannabinoid- containing plant waxes are produced by various plant species, most notably by the flowering cannabis plants from the genus, Cannabis, and family Cannabaceae. The Cannabis genus includes the Cannabis sativa species and the various varieties or strains derived therefrom that are selectively bred to have specific cannabinoid profiles. Other plants and species that are known to produce cannabinoids include rhododendron, licorice, liverwort, Echinacea (Echinacea purpurea and Echinacea angustifolia for example), Acmeiia oleracea and Helichrysum umbraculigerum, for example. Cannabis sativa is cultivated for the production of cannabinoids.

It was found that cannabinoids have a relatively strong physical association with the lipophilic compounds in plant waxes. Mere washing of plant waxes with a suitable solvent for the cannabinoids does not recover the desired fraction of cannabinoids from the solid wax matrix. Extraction with conventional organic solvents used to extract cannabinoids from plants were also found to be ineffective at recovering substantial amounts of cannabinoids from plant waxes. An alternative method of removing entrapped cannabinoids from plant waxes that disrupts the unusually strong association or interactions of the cannabinoids with the plant waxes is required.

Accordingly, a method (100) of extracting cannabinoids from plant waxes is provided and shown in Figure 1 . The method comprises dissolving the plant waxes (101 ) in a solvent having a normal boiling point (bp) above 50 °C and a relative polarity (RP) between 0.1 and 0.4 at an elevated temperature achieved by mild heating, thereby forming a solution of the plant waxes and the cannabinoids contained therein and allowing the solution to cool (103) to precipitate the plant waxes whilst the cannabinoids substantially remain in solution. The waxes need to both melt and substantially dissolve at the elevated temperature to ensure that the entrapped cannabinoids are free to dissolve in the selected solvent. A homogenous solution is obtained at the appropriate elevated or heating temperature after both melting and dissolution has occurred. The plant waxes melt at temperatures above 45 °C, whereas the dissolution of the plant waxes depend strongly on the polarity of the solvent. Solvents with a relative polarity of between 0.1 and 0.4 were determined to be effective in dissolving the plant waxes at an elevated temperature, thereby ensuring that a solution of both the plant waxes and cannabinoids is obtained, whilst also allowing for ease of precipitation of most of the plant waxes at ambient conditions or at a relatively mild cooling temperature.

The method may further include the step of removing or separating the precipitate from the solution (105). The removal of the precipitate may be by any appropriate method including settling, filtration or centrifugation, but preferably filtration. If required, the steps of the method may be repeated on the wax precipitate removed from the cannabinoid solution to extract cannabinoids not extracted in a first extraction process or stage, i.e., any residual cannabinoids that may still be associated with the precipitate. The solvent may be recycled and reused in the method.

The plant waxes are optionally waste waxes obtained after an initial or primary cannabinoid extraction process carried out on a plant. Accordingly, the method may include obtaining the plant waxes from the initial cannabinoid extraction process (107) carried out on plants or plant material including plant waxes. The initial extraction process may be a conventional cannabinoid extraction process with a suitable organic solvent such as ethanol, liquified butane, hexane, methanol, acetone or the like. Conventional cold extraction involves a cooling or winterisation step carried out on the primary extraction solution and results in the precipitation of co-extracted waste waxes which still include significant amounts of cannabinoids. The method accordingly allows for the residual cannabinoids in waste wax to be extracted and provides an efficient separation thereof from the waste wax to allow further processing into cannabinoid-containing products.

The cannabinoids extracted in terms of the method should be as pure as possible. It was determined that the plant waxes should not be heated above 100 °C, preferably not above 60 °C, to minimize or avoid degradation and conversion of the cannabinoids. Melting experiments demonstrated that heating to above 45 °C melted most plant wax components. Substantially all plant waxes melted above 50 °C. The elevated or heating temperature is preferably a temperature between 45 °C and 60 °C. This preferred temperature range for heating is above 45 “C to ensure that the waxes have melted and that solubility of the waxes in the solvent of choice is high enough so that a homogenous solution is obtained. The preferred upper limit is determined by the heat sensitivity of the cannabinoids, which may start to convert or be damaged at temperature above 60 “C. The elevated temperature may also be a temperature between 45 °C and 55 °C, 45 °C and 50 °C, or 55 °C and 60 °C. The elevated temperature may be about 50 °C.

Accordingly, a preferred normal boiling point range for the solvent is between 50 °C and 110 °C, more preferably between 50 °C and 105 °C or 55 °C and 102 °C, or most preferably between 50 °C and 80 °C, or 55 °C and 85 °C. The lower the normal boiling point of the solvent, the lower the cost of solvent recovery in downstream processing.

The relative polarity (RP) of the solvent affects the solubility of the various components of the plant wax. The values for RP are normalized from measurements of solvent shifts of absorption spectra and were extracted from Christian Reichardt, Solvents and Solvent Effects in Organic Chemistry, Wiley-VCH Publishers, 3rd ed., 2003 pp 57-91. Water having an RP of 1 . “Polarity” is a term used to describe the solvation ability of a solvent for charged or neutral, non-polar or dipolar, species. The polarity of a solvent is thus dependent on all possible intermolecular forces between the solvent and the solute molecules, making it impossible to be a measured by an individual physical property (Reichardt, C. & Welton, T., 2010. Empirical Parameters of Solvent Polarity. In: Solvents and Solvent Effects in Organic Chemistry, Wiley, pp. 425-508). RP is a measure of the degree of interaction of the solvent with various polar test solutes. It is therefore an experimentally determined value based on a solvent’s solvation ability. As the methods described herein involve the solubilisation or solvation of wax compounds as the solute, RP is considered the best measure of the required solvation or solubilisation ability of the solvent. Since the RP is an experimentally determined parameter, not all solvents have been tested to date. It will be appreciated by those skilled in the art that further solvents with the appropriate RP may exist and be used in the methods described herein.

Limited data is available on the solubility of plant wax components. The solvent RP range of 0.1 to 0.4 was identified empirically as being suitable for solubilising plant waxes and cannabinoids under mild heating, whilst selectively solubilising just the cannabinoids at near ambient conditions or under mild cooling. Accordingly, selective precipitation of just the plant waxes occurs at or near ambient temperatures, preferably below 30 °C and more preferably at around 20 °C. Solvents with a relative polarity below 0.1 , such as hexane having an RP of 0.01 for example, are unsuitable as they dissolve all the waxes, also at low temperatures, which then prevent the waxes from precipitating out of solution at a preferred temperature range of around 20 °C to 30 °C. Solvents with a relative polarity above 0.4, such as ethanol with an RP of 0.65, resulted in poor purity of the cannabinoid extract due to more wax carry-over.

Differential solubility of the waxes and cannabinoids in a solvent at which precipitation occurs between about 20 °C and about 30 °C was achieved with RPs between 0.1 and 0.4. More particularly, solvent RPs between 0.103 and 0.353 (about 55% of the RP of ethyl acetate) can be set as the RP limits resulting in an optimised and energy-efficient method for extracting cannabinoids from plant waxes. The RP of the solvent may be between 0.103 and 0.353, or 0.1 11 and 0.353, or 0.1 11 and 0.327, or 0.124 and 0.327, or 0.124 and 0.269, or 0.124 and 0.265, or 0.228 and 0.269, or 0.228 and 0.265, for example.

Based on the above boiling point and relative polarity limits, suitable solvents for use in the method of extracting cannabinoids from plant waxes are listed in Table 1 below. A mixture of any of the solvents in Table 1 may also be used.

Table 1 . Physical properties of common solvents that may be used to extract cannabinoids from plant waxes according to the methods described herein.

(Murov S., 2020, Properties of Solvents Used in Organic Chemistry)

The relative polarity of 2-methyl tetrahydrofuran is not available to date. However, it is commonly used to substitute THF and is likely to have a similar RP to THF and within the desired RP range of 0.1 to 0.4. When solvent safety is considered, non-toxic and non-carcinogenic solvents that may be used in the method include 1 ,1 -dichloroethane, 2-butanone, 2-methyl tetrahydrofuran, 3-pentanone, 1 ,2- dimethoxyethane ethyl acetate, methyl acetate, methyl tert-butyl ether or any mixture of these.

To further optimise the purity of the cannabinoid extract, the solvent should substantially lack hydrogen bond donation ability. The hydrogen bond donation ability may be quantified in terms of a Kamlet-Taft acidity parameter (a) approximating 0 (zero). This may render the solvent substantially unreactive towards the cannabinoids or waxes (or both) under conditions of elevated temperature. The elevated temperature may be a temperature obtainable by mild heating. The elevated temperature may be a temperature in a range from about 45 °C to about 60 °C.

It was found that acetone and ethanol, for example, both react with the lipophilic compounds in the plant waxes during the heating step. These reactions are undesirable as they introduce new, potentially problematic compounds or impurities into a cannabinoid production process. Such impurities could affect the downstream distillation conditions and/or reduce the purity of the desired cannabinoid product. The reaction of ethanol and acetone with plant wax components was noted from the appearance of a new liquid phase within the solution that was immiscible with the solvents. It is presumed to be a saponification reaction forming various fatty acid esters of the plant waxes. The filtrate with ethanol and acetone included a significant amount of wax in suspension. This meant a higher degree of carry-over of plant waxes to the separated extract. The solubility of several wax compounds was still high at the 20 °C to 30 °C precipitation conditions tested, resulting in poor product purity.

The suitability of solvents to be used in the method may be further assessed in terms of their Kamlet-Taft parameters for acidity, basicity, and polarity or polarizability (a, p and IT*). The Kamlet-Taft acidity (a) of a solvent quantifies its hydrogen bond donating ability. The Kamlet-Taft basicity (P) quantifies hydrogen bond accepting ability. Both these parameters are indicative of the reactivity of a specific solvent. The values used were reported by Jessop et al (Jessop P.G. et al, Green Chem., 2012,14, 1245-1259) and are summarised in Table 2 below.

Table 2. Kamlet-Taft parameters of commonly used solvents.

Three of the solvents in Table 2 having non-zero a-values were tested for their reactivity towards the wax components in the extract, namely ethanol (a = 0.83), acetone (a = 0.08) and 2-butanone (a =0.06). All three of these solvents illustrated reactivity, which would be undesirable in the extraction process. Accordingly, only the solvents with an acidity or Kamlet-Taft parameter (a) of 0 were considered unreactive such as 1 ,2-dimethoxyethane, 2-methyl tetrahydrofuran, 3- pentanone, cyclopentyl methyl ester, ethyl acetate, methyl acetate and methyl tert-butyl ether.

Preferred non-reactive solvents which optimise the purity of the cannabinoid extract obtained according to method are 1 ,2-dimethoxyethane, 2-methyl tetrahydrofuran, 3-pentanone, ethyl acetate, methyl acetate and methyl tert-butyl ether, most preferably ethyl acetate. Ethyl acetate is a polar apropotic solvent serving only as a medium and not participating in any reactions with the cannabinoids or wax components. Ethyl acetate has a boiling point of 77 °C, a relative polarity of 0.228, a dielectric constant of 6.02, a dipole moment of 1 .88 and a polarity index of 4.4.

The amount of solvent used in the heating step (101 ) aimed at dissolving substantially all the plant waxes may be optimised to extract and recover more cannabinoids. However, in certain embodiments of the method it may be desired to use less solvent, i.e., a lower solvent to wax ratio, in a single extraction and rather repeat the extraction process steps multiple times or in multiple stages or cycles to obtain the same or similar cannabinoid yields. The solvent may also be recycled and reused when the method steps for extracting cannabinoids from plant waxes are carried out more than once. Accordingly, a plant waxes to solvent ratio (g/g) of between about 5 and 20 may be used, or between about 5 and 15, or between about 5 and 1 1 , or between about 6 and 1 1 . A higher solvent to plant waxes ratio (g/g) of for example between about 10 and 20, or about 11 and 20, or about 10 and 15 or about 1 1 and 15, may be used to extract more cannabinoids during a single process cycle or stage.

The heating step (101 ) during which the plant waxes are dissolved in any of the above solvents may be carried out with agitation by stirring for example. The stirring may be stopped during the subsequent cooling step (103) aimed at precipitating most or all of the plant waxes. Since the plant waxes are a mixture of different lipophilic compounds, the various different lipophilic compounds precipitate out of solution at different temperatures. The solution may be allowed to cool passively or actively cooled by removing heat. The solution may be allowed to cool to below 30 °C to precipitate most of the plant waxes, preferably to below 25 °C, more preferably to about 20 °C to precipitate substantially all of the plant waxes. Cooling the solution further will improve the quality of the filtrate by reducing the solubility of the waxes in the solvent even further, allowing more wax compounds to precipitate out. Thus, any temperature lower than 30 “C is suitable and the exact temperature used may depend on other process considerations such as energy efficiency or the like. The solution may, for example, be allowed to cool or cooled to between 0 °C and 30 °C, 10 °C and 20 °C, 20 °C and 30 °C, 5 °C and 25 °C, 5 °C and 15 °C, or 15 °C and 25 °C.

The plant waxes used as feed in the method may be Cannabis plant waxes which are known to contain significant amounts of cannabinoids. The cannabinoids extracted with the method described herein may be further processed to be used in pharmaceuticals to ease or prevent symptoms caused by certain medical conditions or in nutraceuticals. The cannabinoids may therefore be processed to be medical or food grade cannabinoids by having a specific purity and/or by avoiding the use of toxic or carcinogenic solvents in the methods described herein.

The method may be used to extract non-psychoactive or non-psychotropic cannabinoids such as cannabidiol (CBD). Further separation or distillation steps may be required to separate the nonpsychoactive cannabinoids from a cannabinoid mixture that is extracted from the plant waxes according to the methods described herein.

The method is aimed at preserving the acid and/or neutral forms of various cannabinoids to provide material useful for further processing into medical grade or food grade cannabinoid products.

A cannabinoid extract obtained by the method as described above is also provided. The cannabinoid extract includes the solvent used in the methods described herein and is substantially free of plant waxes. The cannabinoid extract contains a significantly higher quantity of cannabinoids in comparison to an extract or solution obtained from conventional solvent washing of the waxes. The cannabinoid extract includes at least about 15% of the original amount of cannabinoids in the plant waxes, preferably at least about 30%, more preferably at least about 60%. The amount of cannabinoids in the extract may depend on the solvent to plant waxes ratio. The solvent in the extract may be evaporated to precipitate the cannabinoids. Alternatively, the extract may be subjected to further separation and purification processes to isolate specific cannabinoid compounds from the mixture extracted.

When the method described above is fully optimised or repeated the amount of cannabinoids that can be extracted from the plant waxes may be more than 80%.

The organic solvents typically used in conventional extraction of cannabinoids from fresh plants or dried plant material, such as ethanol, butane, hexane, methanol and acetone, are not suitable for the extraction of cannabinoids from plant waxes. Plant waxes are a unique feed for the extraction of cannabinoids and the chemical interactions between the cannabinoids and the lipophilic compound of the plant waxes further complicates their extraction. Only solvents having specific properties including a relative polarity of between 0.1 and 0.4 and a normal boiling point above 50 °C were found to be suitable for extracting cannabinoids from plant waxes. The method was developed specifically for plant waxes and validated through testing on plant wax obtained from cannabis plants.

It was found that the method recovers significantly more cannabinoids while using less solvent than cold washing of plant waxes with a solvent. When a selected amount of plant waxes was washed with five times as much solvent as is used in the method described herein, the percentage cannabinoid recovery was found to be only about 30%.

EXAMPLES

Methods

1 . Obtaining cannabis plant wax

Two wax deposits were prepared, wax A originating from a CBD-high strain of cannabis and wax B from a THC-high strain of cannabis. Initial cannabinoid extraction processes were carried out on the respective raw cannabis inflorescences using counter-current flowing ethanol at temperatures ranging between 20 to 24 °C. The extracts were concentrated through evaporation and then cooled to -40 °C during a process called winterisation. At this temperature, the wax precipitates out of solution and is removed through filtration. Both waxes contained residual solvent (ethanol), carried over from filtration, with wax B containing more ethanol than wax A as wax B was not effectively concentrated through evaporation prior to the winterisation step.

2. Cannabinoid extraction from wax A and wax B

A selected amount of wax obtained according to process 1 above, namely 5 g for wax A and 8 g for wax B, was mixed with a selected solvent at amounts such that the ratio of dry wax to solvent was between 1 and 30. Dry wax (solvent free wax) was determined via TGA as described below. To solubilise the cannabinoids, the mixture of solvent and wax were heated to 50“C. The heated sample was removed from the heating source and allowed to cool under ambient conditions (~20 °C), so that the waxes precipitate out of the solution. Cooled samples were subsequently filtered and washed with additional solvent using a Buchner funnel and vacuum pump. The filtrate or extract with high cannabinoid content was collected and the solvent evaporated to yield the recovered cannabinoids.

The above cannabinoid extraction process was repeated using the filtered wax as starting material (stage 2). Figure 2 is a flow diagram of the two-stage process used to extract cannabinoids from the waxes (wax A and wax B, respectively). Tables 2 and 3 include the names and descriptors for the process streams and the units of operation shown in Figure 2.

Table 2. Accompanying stream table for Figure 2. Table 3. Accompanying units of operation table for Figure 2. 3. Cannabinoid quantification

The fraction of cannabinoids present in both wax by-products was quantified through a combination of gravimetric and high-performance liquid chromatography (HPLC) analysis.

3.1 Pre-extraction cannabinoid quantification

A known amount of each of wax A and wax B was dissolved in 100 mL of ethyl acetate. The mixture was cooled to -5 “C, at which point most lipophilic compounds precipitate out of solution. The precipitate was separated from the solution via centrifugation at -5 “C and 4400 RPM for an hour, and the supernatant was drawn off. The supernatant, with substantially all lipophilic compounds removed via centrifugation, was heated until all solvent evaporated and only the cannabinoid fraction remained. This weight was recorded as the mass of cannabinoids present in the wax after corrections were applied to account for entrained cannabinoids remaining in the wax after centrifugation.

3.2 HPLC analysis

The cannabinoid profile was characterised via HPLC. Samples were extracted using an 80:20 (v/v) mixture of methanol and water. The samples were sonicated with an AC-150H sonicator (MRC) for an hour before being centrifuged, after which the supernatant was drawn off and sent for analysis. The analysis was performed on a Synapt G2 instrument from Waters using electrospray ionization. The column used was BEH C18 from Waters with dimensions 2.1 x100 mm and pore size 1.7 pm. The mobile phase used was a mixture of water and acetonitrile. Standards for cannabidiol (CBD), delta-9-tetrahydrocannabinol (A9-THC) and tetrahydrocannabinolic acid (THCA) were available, but (cannabidiolic acid) CBDA was quantified relative to THCA. The highest concentration standard was 33.3 ppm and the samples with higher concentrations were extrapolated.

3.3 Post-extraction cannabinoid quantification

The quantity of cannabinoids present in the extracts obtained according to the process described at point 2 were determined through thermogravimetric analysis (TGA). To prepare the samples for analysis, each sample was placed in a freezer for 30 minutes to precipitate all dissolved waxes. The frozen samples were centrifuged, and the supernatant drawn off and analysed using TGA. The vials containing the wax precipitates from centrifugation were left to dry for over a week and the dried wax weight was recorded to calculate percentage wax carry-over. TGA was performed with a TGA 5500 (Waters) with a gas flowrate of 70 mL/min. The samples were equilibrated at 50 “C for 0.16 min after which it was ramped up to 80 “C at 10 “C/min, where it was maintained at isothermal conditions for two minutes. At this temperature, all solvent was considered evaporated and the remaining mass was determined to be the cannabinoid mass. A further increase in temperature to 350 “C at 20 “C/min evaporates the cannabinoids, with the remaining mass considered to be ash or residual wax.

Results

The above methods were used to extract cannabinoids from wax A and the results obtained with ethyl acetate as extraction solvent are shown in Table 4 below.

Table 4. Experimental results of the method performed on cannabis wax A.

The data indicate that an increase in solvent relative to the dry wax (g/g) improves the recovery of cannabinoids. At a solvent to dry wax ratio (g/g) of 11.3 about 79% (almost 80%) of cannabinoids in the cannabis wax was recovered after repeating the extraction method twice. The method and solvent to wax ratio may be further optimised to obtain recoveries above 80%.

As described herein, the method relies on the dissolution of plant waxes and cannabinoids at an elevated temperature, followed by selective precipitation of just the plant waxes at ambient temperature or a further reduced temperature. At temperatures above 45 °C, preferably about 50 °C near complete dissolution of the plant waxes frees the entrapped cannabinoid fraction and allows it to partition into the solvent phase. Since solubility is highly influenced by the polarity of the solvent, the relative polarity is a critical parameter of solvent selection. It will be appreciated by those skilled in the art that relative polarity is used as an indicator of polarity of the solvent, but that other polarity measures may also be used to quantify the required polarity of the solvent. Precipitation ideally happens at 20 °C, when there is limited dissolution of the waxes in the solvent.

The foregoing description has been presented for the purpose of illustration; it is not intended to be exhaustive or to limit the invention to the precise forms disclosed. Persons skilled in the relevant art can appreciate that many modifications and variations are possible in light of the above disclosure. For example, the solvents listed are examples of solvents having the required properties to be used in the method. All solvents having a relative polarity of between 0.1 and 0.4 and a normal boiling point above 50 °C may be used, with certain solvents being more desirable for being greener (more energy efficient) or safer (non-toxic or non-carcinogenic for example). Furthermore, the reactivity in terms of the acidity of the solvent is only particularly relevant for solvent selection when it is required that the processed waxes be further refined into useable products.

It will further be appreciated that the method, which is aimed at recovering entrained cannabinoids from a wax deposit, may be carried out on any type of plant which produces a wax deposit that includes a desirable quantity of cannabinoids. Furthermore, the presence of a specific cannabinoid in the wax is dependent on the cannabinoid profile of the plant or of a selected Cannabis strain from which the wax is obtained. The method is suitable for extracting any cannabinoid that can be entrained in the wax, as all cannabinoids should be concentrated in the solution or filtrate obtained according to the method. The solution or filtrate may be be returned to an overhead processing stream where it may undergo distillation to isolate specific cannabinoids.

The language used in the specification has been principally selected for readability and instructional purposes, and it may not have been selected to delineate or circumscribe the inventive subject matter. It is therefore intended that the scope of the invention be limited not by this detailed description, but rather by any claims that issue on an application based hereon. Accordingly, the disclosure of the embodiments of the invention is intended to be illustrative, but not limiting, of the scope of the invention, which is set forth in the following claims.

Finally, throughout the specification and accompanying claims, unless the context requires otherwise, the word ‘comprise’ or variations such as ‘comprises’ or ‘comprising’ will be understood to imply the inclusion of a stated integer or group of integers but not the exclusion of any other integer or group of integers.